The cation-independent mannose 6-phosphate receptor (CI-M6PR), a transmembrane glycoprotein of 300 kDa, plays very important roles in many biological processes. The main role of CI-M6PR is transporting and sorting those lysosomal enzymes that contain the mannose 6-phosphate (M6P) recognition marker in their structure from the trans-Golgi network to the lysosomes. CI-M6PR also mediates the endocytosis of extracellular M6P-containing ligands. The M6P-containing proteins, which differ from lysosomal enzymes and are internalized through CI-M6PR transport, include Granzyme B, a protease involved in cytotoxic-T-cell-induced apoptosis; the herpes simplex virus (HSV)7; and even leukemia inhibitory factor (LIF), a multifunctional protein that plays an important role in neuronal, platelet and bone formation. Renin is also internalized by CI-M6PR, which permits its clearance. CI-M6PR also acts on molecules that do not penetrate into cells such as the latent precursor of transforming growth factor-beta (L-TGFβ), the proform of a hormone that regulates cell growth, and acts in another process involved in L-TGFβ activation, the plasminogen/plasmin conversion by a serine protease, the urokinase-type plasminogen activator (uPA). The pro-uPA is proteolytically cleaved and thereby activated when bound at the cell surface to a specific uPA receptor (uPAR) that presents an affinity for CI-M6PR. Moreover, studies suggest that CI-M6PR may act as a tumour suppressor as it modulates the local level of mitogen insulin-like growth factor II (IGF2), and loss of CI-M6PR function is associated with the progression of a high proportion of hepatocarcinomas. As CI-M6PR binds and endocytoses IGF2 in order to decompose it into lysosomal compartments, this receptor is also called M6P/IGF2 receptor. Another ligand of this receptor is retinoic acid, which is involved in apoptosis and growth inhibition. These three ligands (M6P, IGF2, retinoic acid) are recognized by different extracellular binding sites located on CI-M6PR, which contains 15 repeat domains. The phosphate moiety as well as the hydroxyl groups on the mannopyranosidic ring of M6P contribute to a hydrogen-bonding network with two binding sites located on domains 3 and 9 of CI-M6PR. This ability to recognize two M6P residues allows CI-M6PR to bind lysosomal enzymes with high affinity (Kd=2 nM).
The use of this strong affinity between M6P and CI-M6PR has been proposed to develop enzyme replacement therapies, in particular for lysosomal enzymes deficiencies.
However, supplies for the required enzymes are limited and large-scale production of sufficient quantities of enzymes for therapeutic administration is difficult. In addition, lysosomal enzymes purified from recombinant expression systems are often not well phosphorylated and the extent of M6P phosphorylation varies considerably with different enzymes. Lysosomal enzymes lacking in M6P phosphorylation compete poorly for receptor-mediated endocytic uptake by target cells and are thus of limited efficacy in enzyme replacement therapy.
Zhu (U.S. Pat. No. 7,001,994) proposes methods for introducing highly phosphorylated mannopyranosyl oligosaccharides containing M6P to carbonyl groups generated on the glycosidic part of lysosomal enzymes while retaining their biological activity. These mannopyranosyl oligosaccharides containing M6P are typically phosphopentamannose and are chemically treated to contain a carbonyl-reactive group in lieu of an hydroxyl group of the sugar bone. This carbonyl-reactive group is then reacted with an oxidized glycoprotein to form a M6P-glycoprotein. As shown in the experimental section of U.S. Pat. No. 7,001,994, these compounds, whereas conserving a good enzymatic activity, have a poor affinity for the CI-M6PR. According to Zhu, this low affinity is due to steric hindrance of the vicinal aldehyde groups.
These compounds are therefore not suitable for a satisfactory enzyme replacement therapy and there is thus a need for new compounds having a high affinity to the CI-M6PR (thereby allowing a specific addressing of the compounds to the lysosome) and conserving a good biological activity.